Cytokines and growth factors play important roles in regulating cell growth, differentiation and function. The JAK-STAT signaling pathway is the major pathway for transducing signals from a multitude of cytokines and growth factors directly into the nucleus, regulating a diverse array of biological functions including immune response, cellular development, and oncogenesis. The STAT proteins (Signal Transducer and Activator of Transcription) become phosphorylated, dimerize, enter the nucleus and bind to specific DNA sequences to activate transcription. Much analysis has been focused on the transduction of signals by the STAT proteins, while little is known about their mode of stimulating transcription of an otherwise silent gene. To further understand the JAK-STAT pathway, particularly signaling into the nucleus and mechanism of transcription activation by STATs, it is essential to identify the protein players that are brought together by STATs in the process of transcription activation. We will use Stat1 and the IFN-gamma signaling pathway as a model system to test the hypothesis that STAT proteins recruit a set of co-activators, some of which maybe unique to STATs, to regulate gene expression in response to cytokines and growth factors. The central goal of this proposal is to identify the transcription co-activators interacting with Stat1 using affinity-purification and mass spectrometry techniques. The structural and functional importance of these co-activators will be analyzed using biochemical and molecular approaches. Identification of these proteins involved in the process of transcription activation by Stat1 will not only provide insight into the IFN-gamma signaling pathway and other cytokine signaling pathways involving different STATs, but also contribute to the understanding of the general mechanism of transcription activation.